The objectives of the proposed research are four-fold. The first objective is to establish the mechanism whereby NAD ion, cyclic AMP, sugar phosphates, and a heat-stable compound isolated from whole rabbit reticulocyte lysates exert either a stimulatory or inhibitory effect on the initiation step of the protein synthetic process in lysed rabbit reticulocytes and HeLa cell extracts. The second objective involves the role of NAD ion and NAD ion analogs in the nuclear ADP-ribosylation reation. 2'dNAD ion and 3'dNAD ion will be used to elucidate the mechanism by which these two NAD ion analogs restore template restriction in nuclei isolated from Novikoff hepatoma HeLa cells, and fetal rat liver. The role of NAD ion and poly(ADP-ribose) synthase in the DNA repair process of human xeroderma pigmentosum cells as compared to normal human fibroblasts will als be studied. The third objective is to elucidate the precursor for the biosynthesis of the cyclopentenediol ring of base Q in the anticodon region of some species of tRNA and to use this biosynthetic data to determine if the increased Q*tRNA as seen in ascites hepatoma (compared to normal rat liver) is responsible for the transformation of a normal cell into a cancer cell. The final objective is to determine which methylene hydrogen on carbon-4 of L-iso-leucine (i.e., the 4R, 4S, or both hydrogens) is enzymatically removed in the dehydrogenation step in the biosynthesis of 3-ethylidene-L-azetidine-2-carboxylic acid by s. cacaoi. (4R-3H)L-isoleucine will be used for these studies. The retention or loss of tritium will tell us if the 4R and 4S hydrogens are stereochemically nonequivalent in the enzymatic dehydrogenation step.